Indoleamine 2,3-dioxygenase (IDO)

Coptisine has an IC50 value of 6.3 μM and a Ki value of 5.8 μM, making it a very effective non-competitive IDO inhibitor[1]. The growth of A549, H460, H2170, MDA-MB-231, and HT-29 cells is inhibited by copoxine (0.1-100 μM), with IC50 values of 18.09, 29.50, 21.60, 20.15, and 26.60 μM, in that order. In A549 cells, coptisine (12.5, 25, and 50 μM) concentration-dependently causes G2/M arrest and apoptosis, downregulates the expression of cyclin B1, cdc2, and cdc25C, and increases the expression of pH2AX and p21. In A549 cells, copoposite (12.5, 25, 50 μM) likewise causes mitochondrial dysfunction and triggers caspase activation. Additionally, ROS levels are raised by coptisine (50 μM) in a time-dependent manner (0.5, 1, 2, 4, 12, and 24 hours) [3].

Mice's LD50 value for coptisine was 880.18 mg/kg, and its toxicity increased with concentration. The dosage of 154 mg/kg/day for 90 days did not cause toxicity in SD rats. In addition to increasing HDL-c content to varied degrees and slowing down the weight gain brought on by the HFHC diet, copoxine (23.35, 46.7, 70.05 mg/kg, po) increased fecal cholesterol and TBA levels in hamsters in a dose-dependent manner. It also reduced TC, TG, and LDL-c levels in the serum of the animals. Inducing the expression of SREBP-2, LDLR, and CYP7A1 proteins involved in cholesterol metabolism, coptisine (70.05 mg/kg, po) lowers the level of HMGCR protein expression [2].

Recombinant human IDO protein was incubated with different concentrations of Coptisine (0.01-10 μM) and L-tryptophan (substrate) in reaction buffer. After incubation at 37°C for 1 hour, the reaction was terminated, and the concentration of KYN (product of IDO-catalyzed reaction) was measured by high-performance liquid chromatography (HPLC) to calculate IDO enzyme activity inhibition rate and Ki value [1]

BV2 microglial cell experiment: Cells were seeded and stimulated with IFN-γ (20 ng/mL) for 24 hours to induce IDO expression, then treated with Coptisine (0.1-10 μM) for another 24 hours. qPCR was used to detect IDO mRNA expression (with GAPDH as internal control), Western blot to detect IDO protein level, and HPLC to measure KYN and TRP concentrations in cell supernatants [1] - A549 cell proliferation assay: Cells were seeded in 96-well plates and treated with Coptisine (5-40 μM) for 24, 48, 72 hours. MTT reagent was added, and absorbance at 570 nm was measured to calculate cell viability [3] - A549 cell cycle assay: Cells were treated with Coptisine (10-40 μM) for 24 hours, harvested, fixed with ethanol, stained with propidium iodide (PI), and analyzed by flow cytometry to determine cell cycle distribution [3] - A549 cell apoptosis assay: Cells were treated with Coptisine (10-40 μM) for 24 hours, stained with Annexin V-FITC/PI, and apoptotic rate was detected by flow cytometry; mitochondrial membrane potential was measured by JC-1 staining and flow cytometry [3] - Western blot for A549 cells: Cells were treated with Coptisine (10-40 μM) for 24 hours, lysed to extract proteins, separated by SDS-PAGE, transferred to membranes, and probed with antibodies against Bax, Bcl-2, caspase-3, -8, -9, and β-actin. Bands were visualized and quantified [3] - ROS detection in A549 cells: Cells were loaded with DCFH-DA probe, treated with Coptisine (10-40 μM) for 24 hours (some cells pretreated with NAC for 1 hour), and ROS level was detected by flow cytometry [3]

AD mouse model (APP/PS1 double-transgenic mice): 6-month-old male mice were randomly divided into vehicle group and Coptisine treatment groups (20, 40 mg/kg/day, n=10 per group). Coptisine was dissolved in 0.5% carboxymethylcellulose sodium (CMC-Na) solution. Mice received daily oral gavage for 3 months. Cognitive function tests (Morris water maze, novel object recognition, passive avoidance) were performed before sacrifice. Serum, hippocampus, and cortex tissues were collected for KYN/TRP ratio detection, qPCR, Western blot, and immunohistochemistry [1] - Hypercholesterolemic hamster model: Male Syrian golden hamsters were fed a high-fat diet (HFD) for 2 weeks to induce hypercholesterolemia, then randomly divided into HFD control group and Coptisine treatment groups (50, 100 mg/kg/day, n=8 per group). Normal diet group was set as control. Coptisine was dissolved in distilled water and administered by daily oral gavage for 4 weeks. Body weight was recorded weekly. At the end of treatment, serum was collected to detect lipid profiles (TC, TG, LDL-C, HDL-C), and liver tissue was collected for TC/TG content measurement and qPCR analysis [2]

Absorption, Distribution and Excretion
Corydalis saxicola Bunting is an important component of many traditional Chinese medicine formulas. It has been shown to possess various pharmacological activities, including antibacterial, antiviral, and anticancer activities. Its active components include dehydrocarbamide, Coptisine, dehydroapocarbamide, and tetradehydroscoline. This study aimed to investigate the pharmacokinetics and tissue distribution of Corydalis saxicola in rats using high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). The systemic clearance of the four active alkaloids in plasma exceeded 93% of hepatic blood flow, indicating that they are likely rapidly cleared by the liver. After intravenous and oral administration, less than 10% of the drug was excreted in the urine, suggesting that these four alkaloids may undergo significant metabolism in vivo, or that the drug may be excreted via routes other than urine. The excretion of these four alkaloids after oral administration was significantly lower than that after intravenous administration, suggesting a significant first-pass effect after oral administration. The high apparent volume of distribution indicates that these four alkaloids are widely distributed in rats. Our results also indicate that these four alkaloids can be absorbed after oral administration, although less than 15% of the drug is absorbed into the systemic circulation. In summary, the good oral bioavailability of these four active alkaloids in rats makes further research on the extract of Coptis chinensis worthy of improvement in its oral bioavailability.
This study investigated the absorption of Coptisine chloride (COP) and Coptisine erythrin (BRB) as certain traditional Chinese medicine chemical components in human intestinal epithelial cells. Using a Caco-2 (human colon adenocarcinoma cell line) cell monolayer as an intestinal epithelial cell model, the permeability of COP and BRB from the apical (AP side) to the basal lateral (BL side) and from the BL side to the AP side was examined. The concentrations of these two alkaloids were determined using reversed-phase high-performance liquid chromatography-ultraviolet detection. Transport parameters and apparent permeability coefficients (P_app) were calculated and compared with the corresponding parameters for propranolol and atenolol. The P(app) values were also compared with those of previously reported model compounds (propranolol and atenolol). The P(app) values for COP and BRB were (1.103 ± 0.162) × 10⁻⁵ and (1.309 ± 0.102) × 10⁻⁵ cm·s⁻¹ (from AP side to BL side), and (0.300 ± 0.041) × 10⁻⁵ and (1.955 ± 0.055) × 10⁻⁵ cm·s⁻¹ (from BL side to AP side). Their P(app) values were the same as those for propranolol [(2.23 ± 0.10) × 10⁻⁵ cm·s⁻¹] (a transcellular transport marker, used as a high-permeability control). On the other hand, the efflux transport of BRB was 1.49 times higher than the influx transport, with a P(app) value of 0.67. However, the P(app A-->B)/P(app B-->A) value for COP was 3.67, indicating that efflux transport did not participate in its absorption mechanism in the Caco-2 cell monolayer model. Both COP and BRB can be absorbed by intestinal epithelial cells and are completely absorbed compounds. BRB may be involved in the efflux mechanism from the basal side to the apex in the Caco-2 cell monolayer model. To determine the pharmacokinetics, distribution, and interconversion of total alkaloids, Coptisine, coptisine, Coptisine, and palmatine in rats, the total alkaloids and Coptisine were fed to rats, and their contents in plasma, tissues, and the gastrointestinal tract were determined by reversed-phase high-performance liquid chromatography. The peak time of Coptisine in blood was 2.0. The maximum plasma concentrations of Coptisine in rats were 5.0 hours (Cmax 3.7 mg·L⁻¹) and 5.0 hours (Cmax 2.8 mg·L⁻¹), respectively. Coptisine in rat blood can be converted to tetrandrine. After gavage administration of total alkaloids to rats, the gastric Coptisine content decreased monotonically, while the contents of Coptisine, palmatine, and tetrandrine gradually increased, suggesting that Coptisine may be converted to tetrandrine in the stomach. Animal experiments showed that Coptisine and palmatine are mainly distributed in the lungs, followed by the liver; while tetrandrine and Coptisine are mainly distributed in the liver, followed by the lungs. Coptisine can be converted to tetrandrine. The mechanism by which Coptisine reaches two maximum plasma concentrations in blood may be partly related to gastrointestinal propulsion.
Absorption and Transport Mechanisms This study used a Caco-2 cell uptake and transport model to investigate the uptake and transport of Coptisine, palmatine, lutein, and Coptisine. Cyclosporine A and verapamil were added as P-glycoprotein (P-gp) inhibitors, and MK-571 as a multidrug resistance-associated protein 2 (MRP2) inhibitor, respectively. In the uptake experiment, Coptisine, palmatine, lutein, and Coptisine were all uptaken by Caco-2 cells, and their uptake increased in the presence of cyclosporine A or verapamil. In the transport experiment, the apparent P(AP-BL) values of Coptisine, palmatine, lutein, and Coptisine ranged from 0.1 to 1.0 × 10⁶ cm/s, lower than the apparent P(BL-AB) value. The ER values of all compounds were greater than 2. Cyclosporine A and verapamil both increased the uptake of these compounds. P(app) (AP-BL) increased, but P(app) (BL-AB) of Coptisine, palmatine, myristine, and Coptisine decreased; ER value decreased by >50%. MK-571 had no effect on the transmembrane transport of Coptisine, palmatine, myristine, and Coptisine. In the concentration range of 1-100 μM, Coptisine, palmatine, myristine, and Coptisine had no significant effect on the bidirectional transport of Rho123. Coptisine, palmatine, myristine, and Coptisine are all substrates of P-gp; in the concentration range of 1-100 μM, Coptisine, palmatine, myristine, and Coptisine had no inhibitory effect on P-gp. Jiaotai Wan (JTW) is an important traditional Chinese medicine formula, composed of Coptis chinensis and Cortex cinnamomi powder. It is a famous traditional Chinese medicine formula for treating insomnia and has been used for hundreds of years. This study aimed to compare the pharmacokinetic characteristics of five protoCoptisine alkaloids (Coptisine, palmatine, Coptisine, epiCoptisine, and palmatine) – the main active components of cinnamon powder – in normal and insomnia-prone rats. We also investigated the pharmacokinetic differences of these five protoCoptisine alkaloids after single and multiple administrations. An insomnia-prone rat model was established by intraperitoneal injection of a single dose of p-chlorophenylalanine (PCPA). The concentrations of the five protoCoptisine alkaloids in rat plasma were quantitatively analyzed using rapid liquid chromatography-tandem mass spectrometry (LC-MS/MS). Plasma samples were collected at different time points, drug concentration-time curves were plotted, pharmacokinetic curves were constructed, and pharmacokinetic parameters were estimated. Student's t-tests were performed using SPSS 17.0 software. In the single-dose normal control group, the absorption of the five protoCoptisine alkaloids was slow, bioavailability was low, and the time to peak concentration was delayed. Following a single oral dose, the Cmax and Tmax of the five components in insomnia rats were significantly different from those in normal rats. After multiple oral doses, the pharmacokinetic parameters of the five proCoptisine alkaloids in insomnia rats changed considerably. Significant differences (p<0.05) were observed in the main pharmacokinetic parameters (such as Cmax and Tmax) between single and multiple oral doses in normal rats. The absorption of the five components in the multiple-dose group was superior to that in the single-dose group in insomnia rats. Notably, the plasma concentration curve in the multiple-dose model group showed three peaks. The pharmacokinetic behavior of the five components was described in this paper. In both the normal and model groups, the pharmacokinetic behavior of multiple doses was significantly different from that of a single dose; regardless of whether a single or multiple dose was administered, the pharmacokinetic behavior of insomnia rats was significantly different from that of normal rats. Multiple doses may increase the absorption of JTW in insomnia rats, thereby improving its bioavailability and exerting a more active therapeutic effect.

Toxicity Summary
Identification and Uses: Coptisine is a cytotoxic alkaloid found in Coptis chinensis and is associated with Coptisine. It has been used in biochemical research and has been tested as an experimental therapy. Human Exposure and Toxicity: Cytotoxicity of Coptisine was assessed on a range of human and mouse cell lines and compared with known antitumor drugs mitoxantrone, doxorubicin (Dx), and cisplatin (CDDP). Coptisine was cytotoxic to LoVo and HT-29 cells, less toxic to L-1210 cells, and showed partial cross-resistance to the mitoxantrone-resistant human colon cancer cell line LoVo/Dx, while no significant cross-resistance was observed to the cisplatin-resistant mouse leukemia cell line L-1210/CDDP. Compared to many cell- or other structure-associated alkaloids, Coptisine selectively inhibits vascular smooth muscle cell proliferation at lower concentrations. Coptisine possesses potential pharmacological activity in lowering cholesterol, possibly by regulating the mRNA and protein expression of key genes involved in cholesterol metabolism, such as LDLR, CYP7A1, and HMGCR. Animal studies: Coptisine is a potent and reversible inhibitor of type A monoamine oxidase. Coptisine inhibits the proliferation of vascular smooth muscle cells. In subchronic toxicity studies, no deaths or morbidity associated with Coptisine treatment were observed. Furthermore, no abnormalities were found in clinical signs, body weight, organ weight, urinalysis, hematological parameters, gross necropsy, or histopathological examination after oral administration of Coptisine to any of the animals.

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Non-human toxicity values
Oral LD50 in mice: 852.12 mg/kg

[1]. The IDO inhibitor coptisine ameliorates cognitive impairment in a mouse model of Alzheimer's disease. J Alzheimers Dis. 2015;43(1):291-302.

[2]. The safety and anti-hypercholesterolemic effect of coptisine in Syrian golden hamsters. Lipids. 2015 Feb;50(2):185-94.

[3]. Coptisine-induced cell cycle arrest at G2/M phase and reactive oxygen species-dependent mitochondria-mediated apoptosis in non-small-cell lung cancer A549 cells. Tumour Biol. 2017 Mar;39(3):1010428317694565.

[4]. Inhibition activity of a traditional Chinese herbal formula Huang-Lian-Jie-Du-Tang and its major components found in its plasma profile on neuraminidase-1. Sci Rep. 2017 Nov 14;7(1):15549.

Coptisine is an alkaloid that functions as a metabolite. It has been reported in Coptis chinensis, Corydalis yanhusuo, and other organisms with relevant data. See also: Sanguisorba officinalis (partial); Cephalotaxus fortunei (partial).

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Therapeutic Uses
/EXPL THER/ Indoleamine 2,3-dioxygenase (IDO) is the first rate-limiting enzyme in the kynurenine pathway (KP) of tryptophan catabolism and has recently been identified as a potential participant in the pathogenesis of Alzheimer's disease (AD). Coptisine is the main pharmacologically active component of the traditional Chinese medicine formula Wulianhe Decoction (OGT), which has the potential to treat AD. Our recent studies have shown that OGT can significantly inhibit the activity of recombinant human IDO, providing clues to the possible mechanism of OGT in treating Alzheimer's disease (AD). Here, we characterize the effect of Coptisine on IDO in an AD mouse model based on its ability to inhibit it. The results showed that Coptisine is a highly potent non-competitive IDO inhibitor with a Ki value of 5.8 μM and an IC50 value of 6.3 μM. In AbetaPP/PS1 transgenic mice, oral administration of Coptisine inhibited the activity of IDO in the blood, reduced the activation of microglia and astrocytes, thereby preventing neuronal loss, reducing the formation of amyloid plaques, and improving cognitive impairment. Pheochromocytoma (PC12) cells induced by β-amyloid peptide 1-42 and interferon-gamma exhibited decreased cell viability and increased IDO activity, while Coptisine treatment increased cell viability by reversing the increase in IDO activity. In summary, our current findings further confirm the key link between IDO, KP, and AD, and demonstrate that Coptisine, as a novel IDO inhibitor, holds promise as a novel drug for the treatment of AD.
/EXPL THER/ Excessive activation of nuclear factor κB receptor activator ligand (RANKL) signaling leads to increased osteoclast formation and bone resorption. Downregulating RANKL expression and its downstream signaling pathways may be an effective treatment for bone loss diseases such as osteoporosis. This study found that Coptisine (an isoquinoline alkaloid extracted from Coptis chinensis) inhibits osteoclastogenesis in vitro. Although Coptisine has been shown to possess various in vitro and in vivo activities, including antipyretic, anti-photooxidative, dehumidifying, detoxifying, analgesic, and anti-inflammatory effects, its effect on osteoclastogenesis has not been reported. Therefore, we evaluated the effect of Coptisine on osteoclastogenesis of osteoblasts and osteoclast precursor cells in vitro. The addition of Coptisine and 10⁻⁸ M 1α,25(OH)₂D₃ significantly inhibited osteoclast formation in a dose-dependent manner in a co-culture system of mouse bone marrow cells and primary osteoblasts. Reverse transcription polymerase chain reaction (RT-PCR) analysis showed that Coptisine inhibited 1α,25(OH)₂D₃-induced RANKL gene expression in osteoblasts and stimulated osteoprotegerin gene expression. Adding Coptisine to bone marrow macrophage (BMM) culture in the early stages significantly inhibited RANKL-induced osteoclast formation, suggesting that it acts on osteoclast precursors and inhibits the RANKL/RANK signaling pathway. Within the RANK signaling pathway, Coptisine inhibits NF-κB p65 phosphorylation, which is regulated by RANKL in BMM. Coptisine also inhibits the expression of the key RANKL-induced transcription factor NFATc1. Furthermore, 10 μM Coptisine significantly inhibited the survival of mature osteoclasts and their lacunar initiation activity in the co-culture system. Therefore, Coptisine has the potential to treat or prevent various skeletal diseases characterized by excessive bone destruction.
/EXPL THER/ Since myocardial infarction is a leading cause of morbidity and mortality worldwide, protecting the heart from ischemic damage is a hot research topic. Coptisine is an isoquinoline alkaloid extracted from the rhizome of Coptis chinensis. This study aimed to elucidate whether Coptisine has cardioprotective effects and explore its potential mechanism of action using a rat model of myocardial infarction (MI). Myocardial infarction was induced in rats by subcutaneous injection of isoproterenol (85 mg/kg, twice at 24-hour intervals). Rats were randomly assigned to 7 groups: (I) normal group; (II) isoproterenol group; (III) isoproterenol + fasudil group; (IV) isoproterenol (ISO) + isosorbide dinitrate (ISDN); and (V-VII) isoproterenol + Coptisine (25, 50, and 100 mg/kg). Cardiac function and myocardial ischemia markers were assessed after myocardial infarction. Rats were pretreated with Coptisine (25, 50, and 100 mg/kg) for 21 days and then subcutaneously injected with isoproterenol (85 mg/kg) at 24-hour intervals on days 20 and 21. The results showed that Coptisine possesses strong antioxidant activity, maintaining cell membrane integrity, improving mitochondrial respiratory dysfunction, reducing cardiomyocyte apoptosis, and inhibiting RhoA/ROCK expression induced by high-dose isoproterenol. Coptisine exhibited cardioprotective effects in a myocardial infarction model and should therefore be considered a novel adjuvant therapy to alleviate myocardial injury. Uncontrolled cell proliferation and vigorous angiogenesis play a crucial role in the growth and metastasis of osteosarcoma. This study explored novel drugs derived from traditional Chinese medicine that effectively inhibited the growth and metastasis of osteosarcoma. Coptisine, the active ingredient of Coptis chinensis, significantly inhibited the proliferation of invasive osteosarcoma cells. Coptisine induced cell cycle arrest at the G0/G1 phase by downregulating the expression of CDK4 and cyclin D1, and effectively inhibited tumor growth in a xenograft mouse model. Coptisine significantly inhibited the migration, invasion, and capillary-like network formation of osteosarcoma cells by reducing the expression of VE-cadherin and integrin β3 and decreasing STAT3 phosphorylation. Coptisine significantly increased red blood cell and hemoglobin levels, while maintaining them within the normal range. It also moderately increased white blood cell and platelet counts. These data suggest that Coptisine possesses strong anti-osteosarcoma activity with extremely low toxicity, making it a potential candidate drug for anti-osteosarcoma treatment. Coptis chinensis has been used for centuries in China and other Asian countries to treat inflammatory diseases. However, little is known about the chemical components of this medicinal plant and the mechanisms of its anti-inflammatory activity. This study shows that Coptisine (the main component of Coptis chinensis) effectively inhibits the protein and mRNA expression of inducible nitric oxide synthase (iNOS) in lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages, thereby inhibiting nitric oxide (NO) production. Coptisine can also inhibit the production of pro-inflammatory cytokines interleukin-1β (IL-1β) and interleukin-6 (IL-6) by inhibiting the expression of cytokine mRNA. Furthermore, Coptisine can inhibit the degradation of nuclear factor-κBα inhibitor (IκBα) and the phosphorylation of extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), p38 mitogen-activated protein kinase (MAPK), and phosphatidylinositol 3-kinase/Akt (PI3K/Akt). Coptisine has no effect on the expression of Toll-like receptor 4 (TLR-4) and myeloid differentiation factor 88 (MyD88), nor on the binding of lipopolysaccharide (LPS) to TLR-4. Coptisine can also inhibit carrageenan-induced paw edema in rats and reduce the release of TNF-α and NO in inflamed rat tissues. These results indicate that Coptisine inhibits LPS-stimulated inflammation by blocking the activation of nuclear factor-κB, MAPK, and PI3K/Akt in macrophages, and could serve as a drug for the prevention and treatment of inflammatory diseases.

C38H28N2O12S

736.7001

417.051

1198398-71-8

Coptisine;3486-66-6

72322

Yellow to orange solid powder

3.5

0

4

0

24

502

0

C1C[N+]2=C(C=C3C=CC4=C(C3=C2)OCO4)C5=CC6=C(C=C51)OCO6.OS(=O)(=O)[O-]

XYHOBCMEDLZUMP-UHFFFAOYSA-N

InChI=1S/C19H14NO4/c1-2-16-19(24-10-21-16)14-8-20-4-3-12-6-17-18(23-9-22-17)7-13(12)15(20)5-11(1)14/h1-2,5-8H,3-4,9-10H2/q+1

5,7,17,19-tetraoxa-13-azoniahexacyclo[11.11.0.02,10.04,8.015,23.016,20]tetracosa-1(13),2,4(8),9,14,16(20),21,23-octaene

Coptisine sulfate; 1198398-71-8; Coptisine (Sulfate); orb1302372;

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: Please store this product in a sealed and protected environment (e.g. under nitrogen), avoid exposure to moisture and light.

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO : ~1 mg/mL (~2.40 mM)

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

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Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

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Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

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Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

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Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

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Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

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 (Please use freshly prepared in vivo formulations for optimal results.)